JOURNAL OF THEWORLD AQUACULTURE SOCIETY

Vol. 45, No. 6December, 2014

doi: 10.1111/jwas.12155

Meat and Bone Meal as an Alternative for Fish Meal in SoybeanMeal-Based Diets for Florida Pompano, Trachinotus carolinus L.

Waldemar Rossi Jr

Department of Wildlife and Fisheries Sciences, Texas A&M University System, College Station,Texas 77843-2258, USA

D. Allen Davis1

School of Fisheries, Aquaculture, and Aquatic Sciences, Auburn University, 203 Swingle Hall,Auburn, Alabama 36849-5419, USA

AbstractAlternative protein feedstuffs are potential surrogates for fish meal in the diet of Florida pompano.

This study was designed to evaluate the replacement of fish meal with meat and bone meal in a soybeanmeal-based diet for Florida pompano. In Trial I (10 wk), the basal diet (FM15, formulated to contain40% crude protein and 10% lipid) containing 15% fish meal was modified by the isonitrogenousreplacement of fish meal with meat and bone meal producing diets with 10 (FM10), 5 (FM05), and0% (FM0) fish meal. In Trial II (8 wk), the FM0 served as the basal diet (Tau 0) and was modified bythe additive supplementation of taurine to produce three test diets containing 0.25 (Tau 0.25), 0.5 (Tau0.50), and 0.75% (Tau 0.75) taurine. In Trial I, reduced weight gain (220.1%), feed efficiency (0.31),and survival (84.5%) were observed in fish fed the FM0 diet (P< 0.05). In Trial II, the supplementationof taurine improved the growth performance, survival, and protein and energy retention efficiencies ofFlorida pompano (P< 0.05). These results indicate that the supplementation of taurine was required inthese feed formulations and that meat and bone meal is a good alternative ingredient for fish meal insoybean meal-based diets for Florida pompano.

Fish meal has historically been the majorprotein ingredient in aquatic feeds. More than68% of global fish meal production was destinedto the production of aquatic feeds in the lastdecade (Tacon and Metian 2008). Fish meal is aprotein dense, high quality feed ingredient withan excellent amino acid profile, high nutrientdigestibility, and lacks anti-nutritional factorsnormally found in plant feedstuffs (Gatlin et al.2007; NRC 2011). However, the steady produc-tion of around 5–7 million m.t. per year (FAO2009), accompanied by an increased demand,has caused fish meal prices to rise. A threefoldincrease in fish meal prices between 2000 and2008 was observed (Tacon and Metian 2008),reinforcing the awareness that the expansionof the aquaculture industry depends on thedevelopment of alternative protein ingredients.

1 Corresponding author.

Soybean meal has been utilized as an alterna-tive for fish meal, and its inclusion in aquaticfeeds has increased. Global production of soy-bean meal was estimated to be approximately190 mmt during 2013–2014 (USDA ForeignAgricultural Service 2014). The high proteincontent and reasonably balanced amino acid pro-file of the meal favor its utilization in aquaticfeeds for marine species including Florida pom-pano, Trachinotus carolinus (Gothreaux et al.2010). Nevertheless, some of the compoundspresent in the soybean meal are either indi-gestible to most fish species or can confer a seriesof adverse effects in the digestive system, limit-ing its utilization in some aquatic feeds (Gatlinet al. 2007). Reduced performance as a conse-quence of high dietary levels of soybean mealhas been observed in Florida pompano (Richeand Williams 2011). Therefore, the evaluation ofother protein feedstuffs is necessary to optimizefurther plant-based diets for this species.

© Copyright by the World Aquaculture Society 2014

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614 ROSSI AND DAVIS

One such group of protein feedstuffs con-sists of terrestrial animal by-products derivedfrom poultry or mammalian livestock such aspoultry by-product meal and meat and bonemeal, respectively. The digestibility and aminoacid availability of both by-products have beenevaluated in Florida pompano (Williams 2008),showing their applicability as alternative proteinsources. The poultry by-product meal has beenevaluated in practical diets for Florida pompano(Rossi and Davis 2012), but to our knowledge,information is lacking on the utilization of meatand bone meal.

Meat and bone meal is rendered from driedmammalian tissue (Hardy and Barrows 2002).Over 2 mmt of meat and bone meal was pro-duced in the United Sates in 2009 (Swisher2010), almost half of the fish meal producedglobally. Meat and bone meal is a valuable pro-tein source as well as a good source of cal-cium, phosphorus, and trace minerals. It has beenreported that the nutritional quality of this ingre-dient may vary according to the type of rawmaterial used and processing methods (Hendrikset al. 2002). However, improvements in process-ing technology and the utilization of raw materi-als of higher nutritive value (e.g., blood) can leadto the production of a more nutritious meat andbone meal.

An important aspect associated with the reduc-tion of dietary fish meal is that some fish specieshave demonstrated a conditional requirementfor dietary taurine (Kim et al. 2005a, 2005b).Because the ability to biosynthesize this organicacid varies in fish (Yokoyama et al. 2001; Gotoet al. 2003; Kim et al. 2008) and plant feed-stuffs do not supply taurine (NRC 2006), itssupplementation may be required in plant-baseddiets. In addition, the taurine content in animalby-products such as meat and bone meal is foundto vary considerably (Sptize et al. 2003).

The objective of this study was twofold: (1)evaluate the possibility of using meat and bonemeal as an alternative for fish meal in a soy-bean meal-based diet for Florida pompano; and(2) explore the potential need for supplemen-tal taurine in a fish meal-free diet for Floridapompano containing soybean meal and meat andbone meal as the primary protein sources.

Material and Methods

Fish

Two separate batches of juvenile pompano(1.73 and 2.63 g mean weight) were obtainedfrom Harbor Branch Oceanographic Institute(Fort Pierce, FL, USA). Fish were loaded intoa hauling tank equipped with a supplementaloxygen supply system and transported to theAlabama Department of Conservation and Natu-ral Resources Marine Resource Division, ClaudePeteet Mariculture Center (CPMC, Gulf Shores,AL, USA). At CPMC, fish were acclimatizedby slowly replacing the hauling water with localbrackish water (12.0 g/L salinity, 6.6 mg/L dis-solved oxygen, and 7.75 pH) and subsequentlystocked into a 5-m3 fiberglass recirculating tankequipped with an independent biological filter,air lift pumps, and supplemental aeration pro-vided by a regenerative air blower and air dif-fusers. Fish remained in this tank for approx-imately 30 d acclimatizing to local conditionsuntil adequate size for the growth trials wasachieved. During this period, fish were fed toapparent satiation with a 40% crude protein (CP)and 12% crude fat commercial diet (EXTR 400,Rangen Inc., Angleton, TX, USA).

Diets

Experimental diets were manufacturedat Auburn University, School of Fisheries,Aquaculture, and Aquatic Sciences, Auburn,Alabama, USA. They were prepared by mixingpreground dry ingredients and menhaden fish oilin a food mixer (Hobart A200FT, Hobart Corp.,Troy, OH, USA) for 15 min. Boiling water wasblended into the mixture to promote appropriateconsistency for pelleting, and subsequently themoist mash from each diet was passed througha 3.5-mm die. Wet diets were then placed intoa forced air drying oven (<45 C) for approxi-mately 24 h attaining a moisture content of lessthan 10%. Dry diets were stored at −20 C. Priorto use, each diet was crumbled and sieved to anappropriate size. All experimental diets were for-mulated to be isonitrogenous and isocaloric, con-taining 40% CP, 10% lipid, and 4995.1 kcal/kggross energy (GE) using soybean meal as the pri-mary and fixed protein source (Tables 3 and 4).

ALTERNATIVE PROTEIN-BASED DIETS FOR F. POMPANO 615

Trial I

A growth trial was conducted over a 10-wkperiod to evaluate the replacement of fish mealwith a meat and bone meal blend on the pro-duction performance and body composition ofFlorida pompano. The compositions of the dietsutilized in Trial I are presented in Table 3. Thebasal diet (FM15) was formulated to contain15% fish meal as the sole animal protein source.A gradual, isonitrogenous replacement of 5% ofthe fish meal in the basal diet with meat and bonemeal originated three test diets containing 10(FM10), 5 (FM05), and 0% (FM0) of fish meal.The test diets were supplemented with methion-ine to compensate for differences in this aminoacid relative to the basal diet.

Trial II

An 8-wk growth trial was conducted to eval-uate the supplementation of taurine to the FM0diet utilized in Trial I in the growth performanceand body composition of Florida pompano. Thecompositions of the diets utilized in Trial II arepresented in Table 4. The FM0 diet from Trial Iwas reformulated with approximately the samecomposition and used as the basal diet (Tau 0).Three test diets were designed by the additivesupplementation of taurine (0.25%) to the basaldiet. The resulting test diets contained supple-mental taurine at 0.25 (Tau 0.25), 0.5 (Tau 0.50),and 0.75% (Tau 0.75). The supplementation ofmethionine to all experimental diets in Trial IIwas performed to maintain equal levels across alldietary treatments in this study.

Experimental Systems and Husbandry

The experiments were conducted in separatesemi-recirculating systems operating inside of agreenhouse. Each system was comprised of 12circular fiberglass culture tanks, reservoir tank,biological filter, water pump, and supplementalaeration supplied by a regenerative blower andair diffusers. The culture tanks of Trials I andII were different in water volume, holding 0.7and 1.0 m3, respectively. Routine systems main-tenance consisted of partial water exchanges andsiphoning of solids as needed. A 14-h light:10-hdark cycle was provided by natural light during

summer, and ventilation was utilized to reducewater temperature.

Water quality parameters were monitored rou-tinely. Dissolved oxygen, temperature, salinity,and pH were monitored twice daily using anYSI 556 multi-probe meter (YSI Inc., YellowSprings, OH, USA). Total ammonia nitrogenwas analyzed on a weekly basis using an ionselective electrode (Orion EA 940, Thermo Elec-tron Corporation, Beverly, MA, USA).

Florida pompano juveniles were manuallygraded before the beginning of each growthtrial and were subsequently stocked in the cul-ture tanks of Trial I and Trial II at a densityof 22 and 25 fish/m3, respectively. The ini-tial weight (mean weight± SD) of the pompanojuveniles in the Trials I and II was 2.99± 0.28 gand 5.87± 0.2 g, respectively. Stocking proce-dure was done by manually counting and weigh-ing fish groups, followed by a 1-min immersionin chloroquine phosphate (Marex, Aquatronics,Oxnard, CA, USA) at approximately 22.0 mg/L,as a prophylactic measure against Amyloodiniumocellatum, followed by a 1-min fresh water bath.This prophylactic treatment was repeated everytime fish were handled. Trial I preceded Trial IIbut fish were not reutilized.

Each of the test diets was randomly assignedto three replicate culture tanks (n= 3). Fish ineach culture tank were fed twice daily at a rateapproaching apparent satiation. Group weighingof all fish in each culture tank was performedevery 2 wk.

Data Acquisition and Analytical Procedures

The parameters used to evaluate the growthperformance of the fish were weight gain(%)= [(final body weight− initial body weight)/(initial body weight× 100)]; feed efficiencyratio= (wet weight gain/dry feed intake);and thermal growth coefficient= [(final bodyweight1/3 − initial body weight1/3) / (number ofdays×water temperature, C)× 100]. Proteinand energy retention efficiencies (%)= {[(finalbody weight×final body protein)− (initial bodyweight × initial body protein)] / [(total proteinintake (g)× 100)]}; energy retention efficiency(%)= {[(final body weight × final body

616 ROSSI AND DAVIS

energy)− (initial body weight × initial bodyenergy)]/[(total energy intake (g)× 100)]},respectively, also were determined in Trial II.

At the end of the experimental period, Floridapompano from each culture tank in Trials Iand II were group weighed. In the case ofTrial II, fish were returned to their respec-tive culture tanks where they remained for 3 dunder normal water quality and feeding condi-tions. Subsequently, blood samples were takenfrom randomly selected fish in each culturetank (5 h postprandial) via caudal venipunc-ture. The serum was separated by centrifuga-tion at 1300 g (Model 228; Fisher Scientific Cen-trific, Dubuque, IA, USA) and stored frozen at−45 C for subsequent amino acid and osmo-lality analyses. All amino acid analyses (maindietary protein ingredients, Tau 0 diet in TrialII, and serum-free taurine) were performed intriplicate by an external laboratory (New JerseyFeed Laboratory, Inc., Trenton, NJ, USA). Totalserum osmolality was measured in triplicateusing 10 μL of serum by dew point depressionusing a vapor pressure osmometer (Wescor 5100C, Logan, UT, USA) as previously described(Roy et al. 2007).

For the whole-body compositional analyses,four representative fish per culture tank in TrialII were euthanized with 200 mg/L of tricainemethanesulfonate (MS-222; Western Chemical,Inc., Ferndale, WA, USA), homogenized in afood processor, and frozen at−60 C, until furtheranalysis. Dry matter was determined by placingrepresentative portions of each sample in an ovenat 105 C until constant weight was obtained. Pro-tein content of whole fish and experimental dietswas determined by the micro-Kjeldahl method(Ma and Zuazaga 1942). GE was analyzed with asemimicro-bomb calorimeter (Model 1425, ParrInstrument Co. Moline, IL, USA). All analyticalprocedures were conducted in triplicate.

Statistical Analyses

All data were analyzed using one-wayANOVA to determine significant (P< 0.05) dif-ferences among treatment means. When signif-icant differences were identified, the Student –Neuman Keul’s multiple – range test was used

to determine significant differences between

Table 1. Water quality parameters during Trial I (10 wk)and Trial II (8 wk).

Trial I Trial IIParameter Mean± SD Mean±SD

Temperature, C 27.9± 1.5 28.7± 1.5Salinity, g/L 16.5± 2.5 28.5± 2.1Dissolved oxygen, mg/L 6.8± 1.1 6.5± 0.9pH 7.7± 0.1 7.7± 0.2Total ammonia nitrogen, mg/L 0.1± 0.1 0.3± 0.3

treatment means. Regression analyses wereused to evaluate the relationship between inde-pendent and dependent variables for selecteddata. The adjusted R2 was calculated as pre-viously described by Kvalseth (1985). AllStatistical analyses were performed using theSAS® software package (SAS Institute Inc.,Cary, NC, USA).

Results

Florida pompano showed complete acclima-tion to the CPMC’s water and culture systemsand accepted both commercial and experimentaldiets with no noticeable palatability problems.The prophylactic preventive treatment proved tobe effective at preventing A. ocellatum infesta-tions during the growth trials. The results forwater quality are presented in Table 1. All mea-sured water quality parameters were maintainedwithin acceptable ranges for Florida pompano(Watanabe 1995; Weirich and Riche 2006) dur-ing both Trials I and II. The difference in salinity(16.5 g/L in Trial I and 28.5 g/L in Trial II) wasdue to natural variability in the water supply.

The proximate composition and amino acidprofile of fish meal, meat and bone meal, andsoybean meal are shown in Table 2. The CPcontent of meat and bone meal (66.5%) washigher than in fish meal (62.8%) and soybeanmeal (47.6%). Relatively higher crude fat andash were observed in fish meal and meat andbone meal, while fiber was relatively higher insoybean meal. Regarding the amino acid pro-file, soybean meal showed the lowest levels oflysine (2.8%), methionine (0.8%), and threonine(1.6%) relative to fish meal (4.9, 1.8, and 2.3%,respectively) and meat and bone meal (4.5, 1.0,and 2.2%, respectively).

ALTERNATIVE PROTEIN-BASED DIETS FOR F. POMPANO 617

Table 2. Composition of the ingredients fish meal (FM),meat and bone meal (MBM), and soybean meal (SBM) usedin the diets of Trials I and II.a

Ingredients FM MBM SBM

Proximate composition % as isMoisture 9.7 5.4 13.4Crude protein 62.8 66.5 47.6Crude fat 9.5 7.9 1.6Fiber 0.3 0.9 2.7Ash 20.2 18.9 5.9

Essential amino acidsArginine 3.9 3.9 3.3Histidine 1.3 2.1 1.2Isoleucine 2.4 1.4 1.9Leucine 4.3 5.4 3.4Lysine 4.9 4.5 2.8Methionine 1.8 1.0 0.8Phenylalanine 2.4 2.9 2.2Threonine 2.3 2.2 1.6Valine 2.7 3.2 1.8

Nonessential amino acidsAlanine 3.8 4.7 1.7Aspartic acid 6.6 5.8 6.6Cysteine 0.5 0.8 0.9Glutamic acid 8.8 7.4 9.6Glycine 5.1 7.8 1.9Serine 1.9 2.5 2.0Tyrosine 2.1 1.8 1.5

aDetermined by New Jersey Feed Laboratory, Inc., Tren-ton, NJ, USA. Analyses were performed in triplicate.

The analyzed composition of the diets evalu-ated in Trial I is presented in Table 3. The CP val-ues ranged from 36.0 to 37.4% and were slightlylower compared to the formulated value of 40%CP. GE ranged from 4803.9 to 4899.5 kcal/kgand was fairly constant among diets. Digestibleprotein (DP) and digestible energy (DE) val-ues of the FM15 and FM0 diets were 31.8 and28.0%, and 3489.4 and 3011.4 kcal/kg, respec-tively. The DP/DE of the FM15 and FM0 dietswas 0.090 and 0.093 g/kcal, respectively. Theanalyzed composition values of CP and GE,along with analyzed values of taurine of the dietsevaluated in Trial II, are presented in Table 4. CPvalues of the diets ranged from 38.0 to 38.9% andwere relatively closer to the formulated value of40% CP. GE values were fairly constant amongdiets ranging from 4899.5 to 4995.1 kcal/kg.Dietary taurine levels ranged from 0 to 0.82%and were very similar to the formulated levels.The analyzed amino acid composition of the

basal diet (Tau 0) utilized in Trial II is presentedin Table 5. The level of lysine in the Tau 0 dietwas 2.15%, methionine was 0.80%, total sulfuramino acids (methionine+ cysteine) was 1.51%,and taurine was 0.01%.

The growth performance and survival ofFlorida pompano are presented in Table 6. InTrial I, the total replacement of fish meal withmeat and bone meal in the FM0 diet showedto affect negatively the growth performanceand survival of the fish. Pompano fed the fishmeal-free diet (FM0) had the lowest weightgain (220.1%) and feed efficiency (0.31), com-pared to the other treatments (P< 0.05). Areduction in survival (from 97.8 to 84.5%)was observed between fish fed the FM15 orFM10 and the FM0 diet (P< 0.05). In Trial II,lowest (P< 0.05) weight gain (211.9%), feedefficiency ratio (0.28), and survival (71.1%)were observed in pompano fed the Tau 0 diet,while no significant differences were observedamong the other treatments. The level of post-prandial serum taurine increased linearly (Adj.R2 = 0.90) following its inclusion level in thediets (Fig. 1). The serum osmolality rangedfrom 318.6 to 329.3 mmol/kg and did not differamong treatments (P> 0.05, Fig. 2).

The whole-body proximate composition ofFlorida pompano fed graded levels of taurinein Trial II is presented in Table 7. Whole-bodymoisture, CP, and GE did not differ in fishfed taurine supplemented diets, while significantdifferences were observed relative to those fedthe Tau 0 diet. Highest whole-body moisture(75.4%), lowest whole-body CP (11.6%), andGE (1290.6 kcal/kg) were observed in Floridapompano fed the Tau 0 diet (P< 0.05).

The thermal growth coefficient, feed effi-ciency, and protein and energy retention eff-iciencies of Florida pompano fed graded levelsof taurine in Trial II are presented in Figures 3, 4,and 5, respectively. A quadratic relationship wasobserved between each of these performanceparameters and dietary taurine. By calculatingthe vertex of the quadratic curve, the taurineconcentration at which maximum fish perfor-mance occurred was estimated. The thermalgrowth coefficient and feed efficiency of Floridapompano were maximized at the taurine level of

618 ROSSI AND DAVIS

Table 3. Composition of diets containing different levels of fish meal and meat and bone meal fed to Florida pompano(2.99± 0.28 g mean initial weight) for 10 wk (Trial I).

Ingredients FM15 FM10 FM05 FM0

% as isMenhaden fish meal special selecta 15.0 10.0 5.0 0.0Meat and bone meal, with bloodb 0.0 4.9 9.8 14.7Soybean mealc 50.0 50.0 50.0 50.0Menhaden fish oila 5.3 5.3 5.2 5.2Corn starchd 5.7 5.6 5.5 5.4Whole wheatd 16.0 16.0 16.0 16.0Corn gluten meale 5.0 5.0 5.0 5.0Lecithin (soy refined)d 1.0 1.0 1.0 1.0ASA mineral premixf 0.3 0.3 0.3 0.3ASA vitamin premixg 0.5 0.5 0.5 0.5Choline chlorided 0.2 0.2 0.2 0.2Stay C 250 mg/kgh 0.1 0.1 0.1 0.1CaPO4

d 1.0 1.2 1.4 1.6dl-methioninei 0.00 0.04 0.08 0.12Analyzed values (dry matter basis) %, otherwise notedCrude proteinj 36.3 37.4 37.1 36.0Gross energy, kcal/kgj 4899.5 4803.9 4803.9 4803.9Digestible protein (DP)k 31.8 – – 28.0Digestible energy (DE), kcal/kgk 3489.4 – – 3011.4DP/DE, g/kcal 0.09 – – 0.09

Dash ( – )= values not determined.aOmega Protein Inc., Reedville, VA, USA.bMeat and bone meal with blood, 65 RDB, Mid-South Milling Company, Inc., Memphis, TN, USA.cDehulled solvent extracted soybean meal, Faithway Feed Co. Inc., Guntersville, AL, USA.dMP Biochemicals Inc., Solon, OH, USA.eGrain Processing Corporation, Muscatine, IA, USA.fASA Premix (g 100/g of premix) : cobalt chloride, 0.004; cupric sulfate pentahydrate, 0.250; ferrous sulfate heptahydrate,

4.0; manganous sulfate anhydrous, 0.650; potassium iodide, 0.067; sodium selenite, 0.010; zinc sulfate heptahydrate, 13.193;and α cellulose, 81.826.

gASA Premix (g/kg Premix): thiamin HCL, 0.5; riboflavin, 8.0; pyridoxine HCl, 5.0; Ca-pantothenate, 20.0; niacin, 40.0;biotin, 0.040; folic acid, 1.80; cyanocobalamin, 0.002; vitamin A acetate (500,000 IU/g), 2.40; vitamin D3 (400,000 IU/g),0.50; dl-α-tocopheryl acetate, 80.0; and α cellulose, 834.258.

hStay C®, (l-ascorbyl-2-polyphosphate 35% Active C), Roche Vitamins Inc., Parsippany, NJ, USA.iAldrich-Sigma, St. Louis, MO, USA.jCrude protein (N× 6.25) was determined by micro-Kjeldahl method (Ma and Zuazaga 1942). Gross energy was determined

in a semimicro-bomb calorimeter (Model 1425, Parr Instrument Co., Moline, IL, USA).kDetermined in a separate set of diets by an indirect method using chromic oxide (1%) as the nondigestible marker. A

separate group of advanced (ca. 150 g mean weight) juvenile Florida pompano was used. Fecal samples were collected 5.5 hafter feeding by the stripping technique (Austreng 1978). Chromic oxide concentrations were determined as described byMcGinnis and Kasting (1964). The absorbance was read on a spectrophotometer (Spectronic Genesys 5, Milton Roy Co.,Rochester, NY, USA) at 540 nm, after colorimetric reaction. Digestibility calculations were done using standard formulas(NRC 2011).

0.61%, while 0.68 and 0.62% of dietary taurinewas necessary to maximize protein and energyretention efficiencies, respectively.

Discussion

This study provides additional informa-tion regarding the utilization of alternativeingredients in the diet of Florida pompano. The

combination of meat and bone meal with soy-bean meal and other plant-derived ingredientsshowed to support adequate growth performanceof Florida pompano when taurine was supple-mented to the diet. The positive responses ingrowth performance observed with the supple-mentation of taurine in the fish meal-free dietparallel with previous observations (Rossi and

ALTERNATIVE PROTEIN-BASED DIETS FOR F. POMPANO 619

Table 4. Composition of the fish meal-free diets containing graded levels of taurine fed to Florida pompano(5.87± 0.20 g mean initial weight) for 8 wk (Trial II).

Ingredients Tau 0 Tau 0.25 Tau 0.50 Tau 0.75

% as isMeat and bone meal with blooda 14.2 14.2 14.2 14.2Soybean mealb 50.0 50.0 50.0 50.0Menhaden fish oilc 5.3 5.3 5.3 5.3Corn starchd 4.0 5.0 4.8 4.8Whole wheatd 16.0 15.0 15.0 15.0Corn gluten meale 6.9 6.7 6.6 6.3Lecithin (soy refined)d 1.5 1.5 1.5 1.5ASA mineral premixf 0.3 0.3 0.3 0.3ASA vitamin premixg 0.5 0.5 0.5 0.5Choline chlorided 0.2 0.2 0.2 0.2Stay C 250 mg/kgh 0.1 0.1 0.1 0.1CaPO4

d 1.0 1.0 1.0 1.0DL-methioninei 0.10 0.10 0.10 0.10Taurinei 0.00 0.25 0.50 0.75Analyzed values (dry matter basis) %, otherwise notedCrude proteinj 38.7 38.0 38.5 38.9Gross energy, kcal/kgj 4995.1 4995.1 4995.1 4899.5Taurinek 0.01 0.25 0.49 0.82

aMeat and bone meal with blood, 65 RDB, Mid-South Milling Company, Inc., Memphis, TN, USA.bDehulled solvent extracted soybean meal, Faithway Feed Co. Inc., Guntersville, AL, USA.cOmega Protein Inc., Reedville, VA, USA.dMP Biochemicals Inc., Solon, OH, USA.eGrain Processing Corporation, Muscatine, IA, USA.fASA Premix (g 100/g of premix): cobalt chloride, 0.004; cupric sulfate pentahydrate, 0.250; ferrous sulfate heptahydrate,

4.0; manganous sulfate anhydrous, 0.650; potassium iodide, 0.067; sodium selenite, 0.010; zinc sulfate heptahydrate, 13.193;and α cellulose, 81.826.

gASA Premix (g/kg Premix): thiamin HCL, 0.5; riboflavin, 8.0; pyridoxine HCl, 5.0; Ca-pantothenate, 20.0; niacin, 40.0;biotin, 0.040; folic acid, 1.80; cyanocobalamin, 0.002; vitamin A acetate (500,000 IU/g), 2.40; vitamin D3 (400,000 IU/g),0.50; dl-α-tocopheryl acetate, 80.0; and α cellulose, 834.258.

hStay C®, (l-ascorbyl-2-polyphosphate 35% Active C), Roche Vitamins Inc., Parsippany, NJ, USA.iAldrich-Sigma, St. Louis, MO, USA.jCrude protein (N× 6.25) was determined by micro-Kjeldahl method (Ma and Zuazaga 1942). Gross energy was determined

in a semimicro-bomb calorimeter (Model 1425, Parr Instrument Co., Moline, IL, USA).kDetermined by New Jersey Feed Laboratory, Inc., Trenton, NJ, USA. Analyses were performed in triplicate.

Davis 2012) and are consistent with the recentestimation of a quantitative requirement fordietary taurine in Florida pompano (Salze et al.2014).

Research has shown that Florida pompanocan grow well when fed diets with reduced fishmeal and increased concentration of plant-basedingredients. Lazo et al. (1998) fed juvenileFlorida pompano a diet containing 30% fishmeal and 51% soybean meal for 7 wk, observingno negative effects on growth and survival.Quintero et al. (2012) evaluated the replacementof fish meal with soybean meal and soy proteinconcentrate over a 16-wk growth trial. Theyconcluded that fish meal can be reduced to 15%

without negative effects on growth and feedefficiency of Florida pompano, whereas furtherinclusion of the soy products in place of fishmeal negatively affected the performance ofthe fish.

The results of Trial I showed that the uti-lization of meat and bone meal in diets withsoybean meal at 50% was effective at reducingthe level of fish meal from 15 to 5% withoutsignificant effects on growth performance andsurvival of Florida pompano. However, the over-all performance of the fish was low relative tothat observed in other studies (Lazo et al. 1998;Riche 2009; Quintero et al. 2012). Althoughdifferent studies preclude direct comparisons,

620 ROSSI AND DAVIS

Table 5. Amino acid composition of the basal diet (Tau0) used in Trial II.a

Amino acid % of dry matter

Arginine 2.76Cysteine 0.71Histidine 1.07Isoleucine 1.43Leucine 3.30Lysine 2.15Methionine 0.80Phenylalanine 1.97Taurine 0.01Threonine 1.58Tyrosine 1.39Valine 1.73

aDetermined by New Jersey Feed Laboratory, Inc., Tren-ton, NJ, USA. Analyses were performed in triplicate.

Table 6. Weight gain, feed efficiency, and survival ofjuvenile Florida pompano reared in Trial I (10 wk) and inTrial II (8 wk).a

TreatmentInitial

weight, gWeightgain, %

Feedefficiency

Survival,%

Trial IFM15 3.52 287.6a 0.46a 97.8a

FM10 3.42 300.2a 0.45a 97.8a

FM05 3.51 298.6a 0.46a 91.1ab

FM0 3.47 220.1b 0.31b 84.5b

P-value 0.940 0.036 0.006 0.008PSE 0.124 17.622 0.025 2.223

Trial IITau 0 5.80 211.9b 0.28b 71.1b

Tau 0.25 5.76 457.2a 0.46a 97.8a

Tau 0.50 5.80 528.1a 0.50a 86.7a

Tau 0.75 5.80 520.4a 0.53a 91.1a

P-value 0.159 0.010 0.004 0.013PSE 0.101 54.005 0.035 4.297

PSE= pooled standard error of treatment means (n= 3).aMean values within each column with different super-

scripts are statistically different (P< 0.05).

these observations indicated potential limitationsin the diets of Trial I.

Different results have been reported whenmeat and bone meal was evaluated as an alter-native for fish meal. Meat and bone mealsuccessfully replaced fish meal in the diet ofchannel catfish, Ictalurus punctatus (Li et al.2002; Hedrick et al. 2005) and sunshine bass,Morone chrysops × Morone saxatilis (Web-ster et al. 1999) but could replace only part

Pr > F = < 0.001Adj. R2 = 0.90

0

500

1,000

1,500

2,000

2,500

0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9

Seru

m t

auri

ne,

μm

ol/L

Taurine, %

Figure 1. Postprandial (5 h) serum taurine levels ofFlorida pompano (5.87± 0.20 g mean initial weight) fedfish meal-free diets with graded levels of taurine for 8 wk(Trial II).

0

50

100

150

200

250

300

350

Tau 0 Tau 0.25 Tau 0.50 Tau 0.75

Osm

olal

ity,

mm

ol/k

g

Treatments

Figure 2. Serum osmolality (5 h postprandial) of Floridapompano (5.87± 0.20 g mean initial weight) fed fishmeal-free diets with graded levels of taurine for 8 wk (TrialII). Error bars represent mean± SE.

of the dietary fish meal in Japanese flounder,Paralichthys olivaceus (Kikuchi et al. 1997),yellow croaker, Pseudosciaena crocea (Ai et al.2006), and African catfish, Clarias gariepinus(Goda et al. 2007). Besides fish species, initialfish size, and other experimental conditions,potential explanations for different responsesmay also include variations in the nutritionalquality of the ingredient.

As with other animal by-products, the nutritivevalue of meat and bone meal can be affectedby variations both in the raw materials usedand in processing conditions during rendering(Skurray and Herbert 1974). Parsons et al.(1997) evaluated the protein and amino acidbioavailability of 14 meat and bone meals inchicks. The authors found substantial variationsin protein efficiency ratio (from 0.61 to 2.89)and amino acid availability (lysine, from 73 to88%; methionine, from 77 to 91%; cysteine,

ALTERNATIVE PROTEIN-BASED DIETS FOR F. POMPANO 621

Table 7. Proximate composition of juvenile Floridapompano (5.87± 0.20 g mean initial weight) reared in TrialII (8 wk).a

Treatment Moisture, %Crude

protein, %Gross

energy, kcal/kg

Initial 71.6 11.6 1792.5Tau 0 75.4a 11.6b 1290.6b

Tau 0.25 71.1b 13.4a 1696.9a

Tau 0.50 70.6b 13.8a 1792.5a

Tau 0.75 72.4b 13.7a 1696.9a

P-value 0.015 0.003 0.001PSE 0.836 0.299 57.521

PSE= pooled standard error of treatment means (n= 3).aMean values within each column with different super-

scripts are statistically different (P< 0.05).

Pr > F = < 0.001Adj. R2 = 0.73

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.0 0.2 0.4 0.6 0.8 1.0

TG

C

Taurine, %

Figure 3. Thermal growth coefficient (TGC) of Floridapompano (5.87± 0.20 g mean initial weight) fed fishmeal-free diets with graded levels of taurine for 8 wk (TrialII). Vertical line indicates dietary taurine (0.61%) for max-imum estimated TGC (0.099). Some data points overlap(n= 3).

from 37 to 72%) among the meat and bonemeals (Parsons et al. 1997). In Trial I, the FM15and FM0 diets differed in terms of DP (31.8 and28.0%, respectively) and DE (3489.4 and 3011.4kcal/kg, respectively). Although this might havecontributed with the lower performance of fishfed the FM0 diet, it did not explain the overallpoor growth performance of the fish in Trial I.

Considering that lysine, methionine, and totalsulfur amino acid content of the diets wereadequate (5.51, 2.05, and 3.87% of dietaryCP, respectively) according to the estimatedrequirements for Florida pompano reportedby Riche and Williams (2011) and Patro et al.(2011), the reduced growth performance of fishfed the FM0 diet in Trial I was not indicativeof potential limitations in these amino acids.

Pr > F = < 0.001Adj. R2 = 0.74

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.00 0.20 0.40 0.60 0.80 1.00

FE

Taurine, %

Figure 4. Feed efficiency (FE) of Florida pompano(5.87± 0.20 g mean initial weight) fed fish meal-free dietswith graded levels of taurine for 8 wk (Trial II). Verticalline indicates dietary taurine (0.61%) for maximum esti-mated FE (0.53). Some data points overlap (n= 3).

Pr > F = < 0.001Adj. R2 = 0.74

Pr > F = < 0.001Adj. R2 = 0.77

0.0

5.0

10.0

15.0

20.0

25.0

0.0 0.2 0.4 0.6 0.8 1.0

Ret

enti

on, %

Taurine, %

Protein Energy

Figure 5. Protein (solid line) and energy (dashed line)retention efficiencies of Florida pompano (5.87± 0.20 gmean initial weight) fed fish meal-free diets with gradedlevels of taurine for 8 wk (Trial II). Solid-vertical lineindicates dietary taurine (0.68%) for maximum esti-mated PR (19.6%). Dashed vertical line indicates dietarytaurine (0.62%) for maximum estimated ER (19.2%).Some data points overlap (n= 3). ER= energy retention;PR= protein retention.

In fact, the deletion of supplemental lysine ormethionine from the diets in our previous study(Rossi and Davis 2012) had no effect in thegrowth performance of Florida pompano, indi-cating they were not limiting. However, whentaurine was not supplemented in the diets, anindication of reduced growth performance andsignificant reductions in whole-body CP and GEwas noticed in that study.

The reference value of taurine in fish meal is0.32% (NRC 2006), whereas that in meat andbone meal is less than 0.1% of ingredient drymatter (Sptize et al. 2003). In addition, taurinecontent is very low or undetectable in soybeanmeal and other plant feedstuffs (Sptize et al.2003; NRC 2006). Taking these values, the

622 ROSSI AND DAVIS

estimated taurine content in the diets of Trial Iwas less than 0.1%. In fact, the analyzed taurinecontent in the Tau 0 diet used in Trial II was0.01%. Therefore, all dietary treatments in TrialI were likely limiting in taurine according tothe estimated requirement for Florida pompanoranging from 0.54 to 0.65% (Salze et al. 2014).

The supplementation of taurine to the fishmeal-free diet in Trial II improved the growthperformance, survival, and protein and energyretention efficiencies of Florida pompano, con-firming that taurine was limiting. Such posi-tive responses to supplemental taurine in thecurrent study corroborates with observations inother species such as Japanese flounder (Kimet al. 2005a, 2005b); yellowtail, Seriola quin-queradiata (Matsunari et al. 2005; Takagi et al.2008); red sea bream, Pagrus major (Goto et al.2001; Takagi et al. 2010); cobia, Rachycentroncanadum (Lunger et al. 2007; Salze et al. 2012);and European sea bass, Dicentrarchus labrax(Martinez et al. 2004).

According to the quadratic models, the thermalgrowth coefficient (Fig. 3) and feed efficiency(Fig. 4) of Florida pompano were maximizedat the dietary taurine level of 0.61%. Similarly,protein and energy retention efficiencies weremaximized when dietary taurine was at 0.68 and0.62%, respectively (Fig. 5). These values arequite consistent with the estimated requirementfor dietary taurine in Florida pompano (Salzeet al. 2014).

Reduced growth performance of fish fed dietslow in taurine is normally associated with lowlevels of this organic acid in plasma (Lungeret al. 2007). In this study, the postprandial serumtaurine of fish fed the Tau 0 diet was low, whereasa linear increase in serum taurine was observedwith its supplementation in the diets (Fig. 1).The observation of a linear relationship (Adj.R2 = 0.90) between dietary and serum taurineindicates a high availability, within this range ofsupplementation, of synthetic taurine to Floridapompano.

Osmoregulation in response to changes inwater ionic concentration is among the mostcommon natural adaptations in fish, and taurinehas been found to be closely associated with thisprocess (Thoroed and Fugelli 1994). Although

the potential participation of taurine in osmor-regulatory processes was not evaluated in ourstudy, no differences in serum osmolality of fishfed graded levels of taurine were found.

This study demonstrated that meat and bonemeal is a feasible alternative ingredient for fishmeal in the diet of Florida pompano. Addi-tionally, in corroboration with the findings ofprevious studies, meeting the minimum dietaryrequirement for taurine is necessary for the opti-mization of alternative protein-based diets forthis species.

Acknowledgments

This research was supported by funds fromthe National Sea Grant College Program ofthe United States Department of Commerce’sNational Oceanic and Atmospheric Administra-tion (NOAA) under NOAA grants NA080AR4170840, administered by Mississippi–AlabamaSea Grant Consortium. Additional support wasprovided by Auburn University and the AlabamaDepartment of Marine Resources at ClaudePeteet Mariculture Center, Gulf Shores, AL,USA. We thank all students for their valuablecontribution to the conduction of this researchand particularly we thank M.Sc. Melanie A.Rhodes for her technical assistance. The assis-tance of Luke Roy on serum osmolality analysiswas equally appreciated. We would also like toextend our appreciation to Dr. Camilo Pohlenzfor revising this manuscript.

Literature CitedAi, Q. H., K. S. Mai, B. P. Tan, W. W. Xu, Q. Y. Duan, H.

M. Ma, and L. Zhang. 2006. Replacement of fish mealby meat and bone meal in diets for large yellow croaker,Pseudosciaena crocea. Aquaculture 260:255–263.

Austreng, E. 1978. Digestibility determination in fish usingchromic oxide marking and analysis of contents fromdifferent segments of gastrointestinal-tract. Aquacul-ture 13:265–272.

United States Department of Agriculture Foreign Agri-cultural Service. 2014. Accessed October 22, 2014, atwww.fas.usda.gov/commodities.

FAO (Food & Agriculture Organization of the UnitedNations). 2009. State of the World Fisheries & Aqua-culture 2008. UNFAO, Rome, Italy.

Gatlin, D. M., F. T. Barrows, P. Brown, K. Dabrowski,T. G. Gaylord, R. W. Hardy, E. Herman, G. Hu,Å. Krogdahl, R. Nelson, K. Overturf, M. Rust,

ALTERNATIVE PROTEIN-BASED DIETS FOR F. POMPANO 623

W. Sealey, D. Skonberg, E. J. Souza, D. Stone,R. Wilson, and E. Wurtele. 2007. Expanding theutilization of sustainable plant products in aquafeeds:a review. Aquaculture Research 38:551–579.

Goda, A. M., E. R. El-Haroun, and M. A. K. Chowdhury.2007. Effect of totally or partially replacing fish meal byalternative protein sources on growth of African catfish,Clarias gariepinus (Burchell, 1822) reared in concretetanks. Aquaculture Research 38:279–287.

Gothreaux, C. T., R. C. Reigh, M. B. Williams, and E.J. Chesney. 2010. Amino acid availability of soybeanmeal for Florida pompano. North American Journal ofAquaculture 72:309–313.

Goto, T., S. Takagi, T. Ichiki, T. Sakai, M. Endo, T.Yoshida, M. Ukawa, and H. Murata. 2001. Studieson the green liver in cultured red sea bream fed lowlevel and non-fish meal diets: relationship betweenhepatic taurine and biliverdin levels. Fisheries Science67:58–63.

Goto, T., T. Matsumoto, S. Murakami, S. Takagi, and F.Hasumi. 2003. Conversion of cysteate into taurine inliver of fish. Fisheries Science 69:216–218.

Hardy, R. W. and F. T. Barrows. 2002. Diet formulationand manufacture. Pages 506–596 in J. E. Halver, editor.Fish nutrition, 3rd edition. Academic Press Inc., SanDiego, CA, USA.

Hedrick, R. L., T. J. Popma, and D. A. Davis. 2005. Pondproduction and fatty acid profiles of the fillets of channelcatfish reared on diets with different protein sources.North American Journal of Aquaculture 67:304–311.

Hendriks, W. H., C. A. Butts, D. V. Thomas, K. A. C.James, P. C. A. Morel, and M. W. A. Verstegen.2002. Nutritional quality and variation of meat and bonemeal. Asian Australasian Journal of Animal Science15:1507–1516.

Kikuchi, K., T. Sato, T. Furuta, I. Sakaguchi, and Y.Deguchi. 1997. Use of meat and bone meal as aprotein source in the diet of juvenile Japanese flounder.Fisheries Science 63:29–32.

Kim, S. K., T. Takeuchi, A. Akimoto, A. Furuita, T.Yamamoto, M. Yokoyama, and Y. Murata. 2005a.Effect of taurine supplemented practical diet on growthperformance and taurine contents in whole body andtissues of juvenile Japanese flounder, Paralichthys oli-vaceus. Fisheries Science 71:627–632.

Kim, S. K., T. Takeuchi, M. Yokoyama, Y. Murata,M. Kaneniwa, and Y. Sakakura. 2005b. Effect ofdietary taurine levels on growth and feeding behaviorof juvenile Japanese flounder, Paralichthys olivaceus.Aquaculture 250:765–774.

Kim, S. K., H. Matsunari, T. Takeuchi, M. Yokoyama, H.Furuita, Y. Murata, and T. Goto. 2008. Comparisonof taurine biosynthesis ability between juveniles ofJapanese flounder and common carp. Amino Acids35:161–168.

Kvalseth, T. O. 1985. Cautionary note about R2. TheAmerican Statistician 39:279–285.

Lazo, J. P., D. A. Davis, and C. R. Arnold. 1998. The effectsof dietary protein level on growth, feed efficiency

and survival of juvenile Florida pompano, Trachinotuscarolinus. Aquaculture 169:225–232.

Li, M. G. H., B. B. Manning, and E. H. Robinson.2002. Comparison of various animal protein sourcesfor growth, feed efficiency, and body composition ofjuvenile channel catfish, Ictalurus punctatus. Journal ofthe World Aquaculture Society 33:489–493.

Lunger, A. N., E. McLean, T. G. Gaylord, D. Kuhn,and S. R. Craig. 2007. Taurine supplementation toalternative dietary proteins used in fish meal replace-ment enhances growth of juvenile cobia, Rachycentroncanadum. Aquaculture 271:401–410.

Ma, T. and G. Zuazaga. 1942. Micro-Kjeldahl determina-tion of nitrogen. A new indicator and an improved rapidmethod. Industrial & Engineering Chemistry AnalyticalEdition 14:280–282.

Martinez, J. B., S. Chatzifotis, P. Divanach, and T.Takeuchi. 2004. Effect of dietary taurine supple-mentation on growth performance and feed selectionof sea bass, Dicentrarchus labrax, fry fed withdemand-feeders. Fisheries Science 70:74–79.

Matsunari, H., T. Takeuchi, M. Takahashi, and K. Mushi-ake. 2005. Effect of dietary taurine supplementationon growth performance of yellowtail juveniles Seriolaquinqueradiata. Fisheries Science 71:1131–1135.

McGinnis, A. and R. Kasting. 1964. Colorimetric analysisof chromic oxide used to study food utilization byphytophagous insects. Journal of Agricultural and FoodChemistry 12:259–262.

NRC National Research Council. 2006. Nutrient require-ments of dogs and cats. National Academy Press, Wash-ington, DC, USA.

NRC National Research Council. 2011. Nutrient require-ments of fish and shrimp. National Academy Press,Washington, DC, USA.

Parsons, C. M., F. Castanon, and Y. Han. 1997. Proteinand amino acid quality of meat and bone meal. PoultryScience 76:361–368.

Patro, B., R. C. Reigh, and M. B. Williams. 2011. Dietarymethionine requirement of Florida pompano. Aqua-culture America 2011, New Orleans, Louisiana, USA– Meeting Abstract 180.

Quintero, H. E., D. A. Davis, and M. A. Rhodes. 2012.Soy protein concentrate as an alternative ingredient inFlorida pompano, Trachinotus carolinus, diets. Journalof Applied Aquaculture 24(3):247–261.

Riche, M. 2009. Evaluation of digestible energy and proteinfor growth and nitrogen retention in juvenile Floridapompano, Trachinotus carolinus. Journal of WorldAquaculture Society 40:45–57.

Riche, M. and T. Williams. 2011. Fish meal replacementwith solvent-extracted soybean meal or soy protein iso-late in a practical diet formulation for Florida pompano,Trachinotus carolinus, L., reared in low salinity. Aqua-culture Nutrition 17:368–379.

Rossi, W. Jr. and D. A. Davis. 2012. Replacement offish meal with poultry by-product meal in the diet ofFlorida pompano, Trachinotus carolinus, L. Aquacul-ture 338-341:160–166.

624 ROSSI AND DAVIS

Roy, L. A., D. A. Davis, I. P. Saoud, and R. P. Henry.2007. Effects of varying levels of aqueous potassiumand magnesium on survival, growth, and respiration ofPacific white shrimp, Litopenaeus vannamei, reared inlow salinity waters. Aquaculture 262:461–469.

Salze, G., E. McLean, and S. R. Graig. 2012. Dietarytaurine enhances growth and digestive enzyme activitiesin larval cobia. Aquaculture 362-363:44–49.

Salze, G., Rhodes, M. and D.A. Davis. 2014. Estima-tion of the quantitative requirement for dietary tau-rine in Florida pompano, Trachinotus carolinus, Juve-niles. Aquaculture America 2014 Seattle, Washington,USA – Meeting Abstract.

Skurray, C. R. and L. S. Herbert. 1974. Batch rendering:influence of raw materials and processing conditions onmeat meal quality. Journal of the Science of Food andAgriculture 25:1071–1079.

Sptize, A. R., D. L. Wong, Q. R. Rogers, and A. J. Fascetti.2003. Taurine concentrations in animal feed ingredi-ents; cooking influences taurine content. Journal of Ani-mal Physiology and Animal Nutrition 87:251–262.

Swisher, K. 2010. Market report 2009: Amidst a domesticslump, the industry looks globally. Render MagazineApril:10-17.

Tacon, A. and M. Metian. 2008. Global overview on theuse of fish meal and fish oil in industrially compoundedaquafeeds: trends and future prospects. Aquaculture285:146–158.

Takagi, S., H. Murata, T. Goto, M. Endo, H. Yamashita,and M. Ukawa. 2008. Taurine is an essential nutri-ent for yellowtail, Seriola quinqueradiata, fed non-fishmeal diets based on soy protein concentrate. Aquacul-ture 280:198–205.

Takagi, S., H. Murata, T. Goto, H. Hatate, M. Endo,H. Yamashita, H. Miyatake, and M. Ukawa. 2010.Necessity of dietary taurine supplementation for pre-venting green liver symptom and improving growth

performance in yearling red sea bream, Pagrus major,fed nonfish meal diets based on soy protein concentrate.Fisheries Science 76:119–130.

Thoroed, S. and K. Fugelli. 1994. Free amino compoundsand cell volume regulation in erythrocytes from differ-ent marine fish species under hypoosmotic conditions:the role of a taurine channel. Journal of ComparativePhysiology B164:1–10.

Watanabe, W. O. 1995. Aquaculture of the Florida pompanoand other jacks (Family Carangidae) in the westernAtlantic, Gulf of Mexico and Caribbean basin: statusand potential. Pages 185–205 in K. L. Main and C.Rosenfeld, editors. Culture of high-value marine fishes.Oceanic Institute, Honolulu, HI, USA.

Webster, C. D., L. G. Tiu, A. M. Morgan, and A. Gannam.1999. Effect of partial and total replacement of fishmeal on growth and body composition of sunshinebass, Morone chrysops×M. saxatilis fed practical diets.Journal of World Aquaculture Society 30:443–453.

Weirich, C. and M. Riche. 2006. Acute tolerance of juvenileFlorida pompano, Trachinotus carolinus L., to ammoniaand nitrite at various salinities. Aquaculture Research37:855–861.

Williams, T. 2008. An assessment of alternative feed ingre-dients in practical diets for Florida pompano, Trachino-tus carolinus, held in low salinity recirculating sys-tems. Master’s thesis. The University of Maine, Orono,Maine, USA.

Yokoyama, M., T. Takeuchi, G. Park, and J. Nakazoe.2001. Hepatic cysteinesulphinate decarboxylase activ-ity in fish. Aquaculture Research 32:216–220.